title: Maturation, not initiation, is the major roadblock during reprogramming toward pluripotency from human fibroblasts.
abstract: Pluripotency can be induced in somatic cells by forced expression of POU domain, class 5, transcription factor 1 (OCT3/4), sex determining region Y-box 2 (SOX2), Kruppel-like factor 4 (KLF4), myelocytomatosis oncogene (c-MYC) (OSKM). However, factor-mediated direct reprogramming is generally regarded as an inefficient and stochastic event. Contrary to this notion, we herein demonstrate that most human adult dermal fibroblasts initiated the reprogramming process on receiving the OSKM transgenes. Within 7 d, ~20% of these transduced cells became positive for the TRA-1-60 antigen, one of the most specific markers of human pluripotent stem cells. However, only a small portion (~1%) of these nascent reprogrammed cells resulted in colonies of induced pluripotent stem cells after replating. We found that many of the TRA-1-60-positive cells turned back to be negative again during the subsequent culture. Among the factors that have previously been reported to enhance direct reprogramming, LIN28, but not Nanog homeobox (NANOG), Cyclin D1, or p53 shRNA, significantly inhibited the reversion of reprogramming. These data demonstrate that maturation, and not initiation, is the limiting step during the direct reprogramming of human fibroblasts toward pluripotency and that each proreprogramming factor has a different mode of action.
authors.affiliation: Department of Reprogramming Science, Center for iPS Cell Research and Application, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Koji
authors.initials: K
authors.lastname: Tanabe
authors.forename: Michiko
authors.initials: M
authors.lastname: Nakamura
authors.forename: Megumi
authors.initials: M
authors.lastname: Narita
authors.forename: Kazutoshi
authors.initials: K
authors.lastname: Takahashi
authors.forename: Shinya
authors.initials: S
authors.lastname: Yamanaka
mesh_terms: D002454:Cell Differentiation; D005347:Fibroblasts / Q000166:cytology*; D006801:Humans; D000090062:Kruppel-Like Factor 4; D039904:Pluripotent Stem Cells / Q000166:cytology*
keywords: ES cells; HDF; iPS cell; stemness; transcription factor